01991naa a2200277 a 450000100080000000500110000800800410001902400460006010000160010624501060012226000090022850000430023752011200028065000240140065300310142465300220145565300090147770000200148670000230150670000150152970000250154470000200156970000220158970000220161177300800163321680592024-10-14       2024    bl uuuu     u00u1 u    #d7 ahttps://doi.org/10.1093/jimb/kuae0282DOI1 aROSA, J. da  aOptimizing dsRNA engineering strategies and production in E. coli HT115 (DE3).h[electronic resource]  c2024  aNa publicação: Elibio Leopoldo Rech.  aProducing double-stranded RNA (dsRNA) represents a bottleneck for the adoption of RNA interference technology in agriculture, and the main hurdles are related to increases in dsRNA yield, production efficiency, and purity. Therefore, this study aimed to optimize dsRNA production in E. coli HT115 (DE3) using an in vivo system. To this end, we designed a new vector, pCloneVR_2, which resulted in the efficient production of dsRNA in E. coli HT115 (DE3). We performed optimizations in the culture medium and expression inducer in the fermentation of E. coli HT115 (DE3) for the production of dsRNA. Notably, the variable that had the greatest effect on dsRNA yield was cultivation in TB medium, which resulted in a 118% increase in yield. Furthermore, lactose induction (6 g/L) yielded 10 times more than IPTG. Additionally, our optimized up-scaled protocol of the TRIzol™ extraction method was efficient for obtaining high-quality and pure dsRNA. Finally, our optimized protocol achieved an average yield of 53.3 µg/mL after the production and purification of different dsRNAs, reducing production costs by 72%.  aDouble-stranded RNA  aBacterial dsRNA production  aLactose induction  aRNAi1 aVIANA, A. J. C.1 aFERREIRA, F. R. A.1 aKOLTUN, A.1 aMERTZ-HENNING, L. M.1 aMARIN, S. R. R.1 aRECH FILHO, E. L.1 aNEPOMUCENO, A. L.  tJournal of Industrial Microbiology and Biotechnologygv. 51, kuae028, 2024.